Pages 1-9Review Article
Noriyuki Matsuda and Keiji TanakaDoes Impairment of the Ubiquitin-Proteasome System or the Autophagy-Lysosome Pathway Predispose Individuals to Neurodegenerative Disorders such as Parkinson’s Disease?Abstract: About twenty years ago, an abnormal enrichment of ubiquitin in the inclusion bodies of various neurodegenerative disorders was reported. To date, this phenotype has been a diagnostic feature of many neurodegenerative disorders including Alzheimer’s and Parkinson’s diseases (PD). Because ubiquitin tags proteins that must be eliminated from cells, thereby targeting them for proteasomal degradation, many scientists believed that the ubiquitin-proteasome system (UPS) was inactivated in these neurodegenerative disorders. This inactivation would lead to an accumulation of ubiquitylated proteins with their concomitant aggregation into inclusion bodies and subsequent neuronal death. This hypothesis was further fuelled by the discovery that parkin, the causal gene of autosomal recessive juvenile Parkinsonism, functions as a ubiquitin ligase. However, recent findings by several groups demonstrated that ubiquitylation is also relevant to the autophagy system, with parkin promoting autophagy of dysfunctional mitochondria following the loss of mitochondrial membrane potential. These novel topics do not necessarily mean that the proteasome is involved in neurodegeneration of PD. In this review, we describe current evidence and controversies regarding the relationship between UPS and neurodegenerative disorders such as PD, and discuss several scientific discrepancies that await further clarification.

Pages 11-26Review Article
Diego Albani, Letizia Polito, Gianluigi Forloni (Communicated by Alessandro Serretti)Sirtuins as Novel Targets for Alzheimer’s Disease and Other Neurodegenerative Disorders: Experimental and Genetic Evidence Abstract: Sirtuins are a family of conserved proteins with deacetylase and ADP-ribosyltransferase activity. In humans they are coded by seven genes (SIRT1-7). The most widely investigated and best known sirtuin is SIRT1,whichcan be activated by the natural phytocompound resveratrol and plays a role in several physiologic (embryogenesis, glucose metabolism, apoptosis, autophagy, chromatin integrity, and transcriptional state) and pathologic (diabetes, cancer, cardiovascular disorders, and neurodegeneration) conditions. In the field of neurodegeneration, resveratrol and SIRT1 have proved beneficial in in vitro and in vivo models of Alzheimer’s disease (AD), reducing amyloid-β protein accumulation, considered one of the pathogenic mechanisms. In contrast to these promising biological data, however, genetic studies linking SIRT1 variability to AD are negative (this is the case for other sirtuins too, e.g., SIRT3). In this review, we summarize the in vitro, in vivo, and genetic experimental results linking SIRT1 and the other sirtuins to AD, while a description of sirtuins’ biochemical features and modulating compounds, as well as sirtuins’ involvement in other neurodegenerative disorders are discussed as collateral aims.

Pages 31-35Short Communication
Pei-Jing Cui*, Lan Zheng*, Li Cao, Ying Wang, Yu-Lei Deng, Gang Wang, Wei Xu, Hui-Dong Tang, Jian-Fang Ma, Ting Zhang, Jian-Qing Ding, Qi Cheng, Sheng-Di Chen (Communicated by Thomas Ala) *contributed equally to this work. CALHM1 P86L Polymorphism is a Risk Factor for Alzheimer’s Disease in the Chinese PopulationAbstract: We conducted a case-control study to determine the prevalence of CALHM1 P86L polymorphism (rs2986017) in patients with Alzheimer’s disease (AD) in the Chinese population of mainland China, and also to clarify whether this polymorphism is a risk factor for AD. Fourteen heterozygous P86L carriers were identified among 198 AD patients. One control subject was also found to be a P86L heterozygous carrier. The allelic frequencies of the AD patients and control subjects were found to be significantly different. Our study indicates that the CALHM1-P86L polymorphism is associated with AD in the ethnic Chinese Han.

Pages 37-56Gema Huesa, María Antonia Baltrons, Pilar Gómez-Ramos, Asunción Morán, Agustina García, Juan Hidalgo, Silvia Francés, Gabriel Santpere, Isidre Ferrer, Elena GaleaAltered Distribution of RhoA in Alzheimer’s Disease and AβPP Overexpressing Mice Abstract: RhoGTPases control cytoskeleton dynamics thereby modulating synaptic plasticity. Because Alzheimer’s disease (AD) is characterized by synaptic dysfunction, we sought to determine whether the expression, activity, or localization of the GTPases RhoA, Rac1 and Cdc42, as well as p21-PAK, a downstream target of Rac1/Cdc42, were altered in 18-month-old AβPP Tg2576 mice (Swedish mutation) or in brains from patients with AD and, for comparison in the case of RhoA, Pick’s disease (PiD), a neurodegenerative disorder characterized by hyper-phosphorylated tau accumulation. Immunohistochemical analyses revealed a distinct localization of each RhoGTPase in synapses, dendrite shafts, neuronal bodies, or astrocytes. The association of RhoA with synapses and dendritic microtubules was confirmed by electron microscopy. In AβPP mice, RhoA expression decreased in synapses and increased in dystrophic neurites, suggesting altered subcellular targeting of RhoA. In AD, RhoA immunostaining decreased in the neuropil and markedly increased in neurons, co-localizing with hyperphosphorylated tau inclusions, as though RhoA were sequestered by neurofibrillary tangles. Additionally, total RhoA protein was lower in the AD brain hippocampus, reflecting loss of the membrane bound, presumably active, GTPase. RhoA colocalized with hyperphosphorylated tau in PiD, again suggesting that altered subcellular targeting of RhoA is related to neurodegeneration. No major changes immunohistochemical changes were observed for Rac1, Cdc42, or p21-PAK, thus identifying RhoA among RhoGTPases as a possible therapeutic target in AD.

Pages 57-68Chiara Cupidi, Raffaella Capobianco, Donato Goffredo, Gabriella Marcon, Bernardino Ghetti, Orso Bugiani, Fabrizio Tagliavini, Giorgio GiacconeNeocortical Variation of Aβ Load in Fully Expressed, Pure Alzheimer’s DiseaseAbstract: The relationship between amyloid-β (Aβ) deposition and tau-related neurofibrillary changes is a key issue in the pathogenesis of Alzheimer’s disease (AD). The aim of this study was to investigate the extent and cortical distribution of Aβ and tau pathology, their mutual links and their correlation with the duration of the disease in thirty-nine patients with fully expressed AD. By tau immunohistochemistry, we identified different patterns of distribution of neurofibrillary changes that were ascribed to Braak stage V and VI. The disease duration was longer in patients at Braak stage VI than in those at V. Morphometric analysis carried out in several neocortical areas demonstrated that Aβ load was not uniform among individuals and also varied in the same patient throughout the neocortex, showing decreased severity from associative fields in the premotor and primary motor areas. Aβ load was higher at Braak stage VI than at stage V and correlated positively with disease duration in primary motor cortex and in superior temporal gyrus. Overall, we documented a marked heterogeneity in the extent of Aβ deposition even in AD brains at final stages of disease that cannot be completely explained by a simple, regular build up of this pathologic protein in the cerebral cortex during the course of the disease. This study may be relevant for the correct evaluation of therapeutic strategies for AD that specifically address Aβ pathology.

Pages 69-77Erin L. Heinzen, Anna C. Need, Kathleen M. Hayden, Ornit Chiba-Falek, Allen D. Roses, Warren J. Strittmatter, James R. Burke, Christine M. Hulette, Kathleen A. Welsh-Bohmer, David B. GoldsteinGenome-Wide Scan of Copy Number Variation in Late-Onset Alzheimer’s DiseaseAbstract: Alzheimer’s disease is a complex and progressive neurodegenerative disease leading to loss of memory, cognitive impairment, and ultimately death. To date, six large-scale genome-wide association studies have been conducted to identify SNPs that influence disease predisposition. These studies have confirmed the well-known APOE ε4 risk allele, identified a novel variant that influences disease risk within the APOE ε4 population, found a SNP that modifies the age of disease onset, as well as reported the first sex-linked susceptibility variant. Here we report a genome-wide scan of Alzheimer’s disease in a set of 331 cases and 368 controls, extending analyses for the first time to include assessments of copy number variation. In line with previous reports, no new SNPs show genome-wide significance. We also screened for effects of copy number variation, and while nothing was significant, a duplication in CHRNA7 appears interesting enough to warrant further investigation.

Pages 79-95Ayelén Bulloj*, María C. Leal*, Huaxi Xu, Eduardo M. Castaño, Laura Morelli *contributed equally to this workInsulin-Degrading Enzyme Sorting in Exosomes: A Secretory Pathway for a Key Brain Amyloid-β Degrading Protease Abstract: The accumulation of amyloid-β (Aβ) peptides in senile plaques is one of the hallmarks of Alzheimer’s disease (AD) progression. The endocytic pathway has been proposed as a major subcellular site for Aβ generation while the compartments in which Aβ-degrading proteases interact with Aβ are still elusive. It was suggested that extracellular Aβ degradation may take place by plasma-membrane associated proteases or by extracellular proteases, among which insulin-degrading enzyme (IDE) is the most relevant. However, the mechanisms of IDE secretion are poorly understood. In the present study we used N2a cells to explore if IDE is indeed released through exosomes and the effect of exosomes release on extracellular levels of Aβ. We demonstrated that proteolytically-active plasma membrane associated-IDE is routed in living N2a cells to multivesicular bodies and subsequently, a major fraction is sorted to exosomes. We described that extracellular IDE levels decrease if the generation of multivesicular bodies is interfered with and may be positively modulated by exosomes release under stress-induced conditions. Our results reinforce the relevance of functional IDE in the catabolism of extracellular Aβ.

Pages 97-109Robert K. Fujimura, Teresita Reiner, Fangchao Ma, Virginia Phillips, Alicia de las Pozas, Dennis W. Dickson, Bernard A. Roos, Guy A. Howard, Carlos Perez-StableChanges in the Expression of Genes Associated with Intraneuronal Amyloid-β and Tau in Alzheimer’s Disease Abstract: The clinical hallmark of Alzheimer’s disease (AD) is impairment of cognition associated with loss of synapses, accumulation of amyloid-β (Aβ) both within neurons and as extracellular deposits, and neurofibrillary degeneration composed of phospho-tau. Neurons in the hippocampus are among those that are most vulnerable. The purpose of this study was to investigate the expression of genes associated with cognition, synapse, and mitochondrial function in hippocampal neurons of AD compared to normal individuals. Neurons from the hippocampus with intraneuronal Aβ immunoreactivity were captured with laser microdissection; RNA was extracted; and levels of brain-derived neurotrophic factor (BDNF), TrkB (BDNF receptor), dynamin-1 (DYN), and cytochrome C oxidase subunit II (COX2) were assessed with quantitative real time-polymerase chain reaction. We found no significant differences in the expression of these genes in AD between neurons associated with Aβ compared to those lacking Aβ immunoreactivity. Double immunofluorescence microscopy showed the number of hippocampal neurons coexpressing Aβ or phospho-tau and either BDNF, TrkB, or DYN was similar in AD and controls. Our results suggest that intraneuronal Aβ or phospho-tau do not have obligatory effects on reducing the expression of genes important for memory and cognition in hippocampus of AD.

Pages 111-116Barbara Borroni, Antonella Alberici, Mario Grassi, Marinella Turla, Orazio Zanetti, Angelo Bianchetti, Giorgio Dalla Volta, Renzo Rozzini, Nicola Gilberti, Giuseppe Bellelli, Alessandro PadovaniIs Frontotemporal Lobar Degeneration a Rare Disorder? Evidence from a Preliminary Study in Brescia County, ItalyAbstract: Frontotemporal Lobar Degeneration (FTLD) has always been considered a rare disorder, but only a few epidemiologic studies are available. The aim of the present work was to ascertain all FTLD patients in a Northern Italy area from January 2001 to December 2008, and to estimate the disease prevalence. On the census day, 213 FTD patients were still alive, resulting in an overall prevalence of 17.6 per 100,000 inhabitants. The prevalence of FTLD in patients aged 45-65 years was 22 per 100,000 inhabitants (95% CI=17-27). The prevalence of FTLD was the highest in patients aged 66-75 (78 per 100,000 inhabitants, 95% CI=56-100), and it was still high over 75 years (54 per 100,000 inhabitants, 95% CI=36-69). FTLD is a more common form of dementia than previously recognized. Our results claimed that FTLD is not only an early-onset disorder, but it is frequent in advanced age as well.

Pages 117-127Tim Vanmierlo, Vincent W. Bloks, Leonie C. van Vark-van der Zee, Kris Rutten, Anja Kerksiek, Silvia Friedrichs, Eric Sijbrands, Harry W. Steinbusch, Folkert Kuipers, Dieter Lütjohann, Monique Mulder (Communicated by Robert Friedland)Alterations in Brain Cholesterol Metabolism in the APPSLxPS1mut mouse, a Model for Alzheimer’s DiseaseAbstract: Disturbances in cerebral cholesterol metabolism have been implicated in the pathogenesis of Alzheimer’s disease (AD). Here, we provide evidence that alterations in brain cholesterol homeostasis also can be a consequence of disease progression. We found that APPSLxPS1mut mice, at the age of 9 months when AD-like pathology starts to develop, display increased levels of the cholesterol precursor desmosterol and of the cholesterol metabolite 27-hydroxy(OH) cholesterol in their cerebellum in comparison with wild-type controls. At the age of 21 months, when APPSLxPS1mut brain contains abundant amyloid deposits, desmosterol levels had further increased (>200% in comparison with wild-type mice) in all brain regions examined. 24(S)-OHcholesterol levels were increased in hippocampus and cerebellum of the APPSLxPS1mut mice, while 27-OHcholesterol levels were increased in cerebellum exclusively. Brain cholesterol levels remained unaffected. In line with the fact that desmosterol and 24(S)-OHcholesterol are Liver X Receptor (LXR) activators, the LXR-target genes Abca1 and Apoc1 were upregulated predominantly in hippocampus of APPSLxPS1mut mice at both ages evaluated. The reduced expression of the enzyme that converts desmosterol into cholesterol, the Selective AD indicator 1 gene (Seladin-1/Dhcr24), in both cortex and cerebellum may underlie the increased desmosterol levels in 21 month-old APPSLxPS1mut mice.

Pages 129-135Hypothesis
Adnan ErolAre Paradoxical Cell Cycle Activities in Neurons and Glia Related to the Metabolic Theory of Alzheimer’s Disease?Abstract: The progression and outcome of neurological diseases are determined by the balance between neuron destruction, neuroprotection, and regeneration. In this context, astroglial cells are invariably involved in every kind of neuropathology. Mitotically, active glial cells provide metabolic support to active neurons, contribute to coupling between synaptic activity and local blood flow, and thus protect against oxidative stress. Disturbances of the complex neuron-glia interrelation are increasingly recognized as a potentially important pathophysiological mechanism in a wide variety of neurological disorders including those marked by neurodegeneration. Peripheral insulin resistance-mediated increased oxidative stress in glial cells, and consequent DNA damage, induces senescence in glial cells leads to the development of an inflammatory environment. The immune mediators released by senescent (activated) glial cells are considered to be neurotoxic and ultimately increase the oxidant load of neurons. While the neuron is viewed as the prototypical post-mitotic, fully differentiated cell, certain subsets of neurons reactivate cell-cycle activity in response to triggers of neuronal apoptosis, such as genotoxic stress generated by redox changes due to pathological alterations in supporting astroglial cells. Thus, a paradoxical cell cycle block in glial cells coupled with concomitant cell cycle re-entry in neurons (due to pathological alterations created by peripheral insulin resistance-induced neuroendocrine signaling changes) may cause neurodegeneration, such as seen in Alzheimer’s disease.

Pages 137-145Teresa Nunes, Isabel Fragata, Filipa Ribeiro, Teresa Palma, João Maroco, Jorge Cannas, Mário Secca, Cristina Menezes, Isabel Carmo, Gil Cunha, Miguel Castelo Branco, Manuela Guerreiro, Alexandre de MendonçaThe Outcome of Elderly Patients with Cognitive Complaints but Normal Neuropsychological TestsAbstract: Elderly patients may present with prominent cognitive complaints and have performances in neuropsychological tests within the normal range for the age and education, and thus do not fulfill the criteria for mild cognitive impairment (MCI). There is insufficient evidence to support the clinical decision in these cases (“pre-MCI”). Forty-three subjects, 11 controls, 15 “pre-MCI,” and 17 MCI, were followed for about three and half years with neuropsychological testing and magnetic resonance imaging including volumetric measurements of the hippocampus and amygdala. Two of the “pre-MCI” subjects suffered cognitive and functional deterioration and were diagnosed with dementia. Although the “pre-MCI” subjects as a group had no significant deterioration in neuropsychological tests, they suffered a decline in the total hippocampal volume (P=0.04) along the follow-up time. In contrast, all control subjects remained stable and had no volumetric decreases. As expected, MCI patients underwent significant deterioration in several neuropsychological tests, often progressed to Alzheimer’s disease, and showed decreases both in total hippocampal and amygdalar volumes. Elderly people presenting with cognitive complaints may be in an initial phase of a degenerative disorder and should be followed clinically, even if they have normal neuropsychological tests.

Pages 147-159Laura Serra, Mara Cercignani, Delia Lenzi, Roberta Perri, Lucia Fadda, Carlo Caltagirone, Emiliano Macaluso, Marco BozzaliGrey and White Matter Changes at Different Stages of Alzheimer’s DiseaseAbstract: This study investigates abnormalities of grey (GM) and white matter (WM) in Alzheimer’s disease (AD), by modeling the AD pathological process as a continuous course between normal aging and fully developed dementia, with amnesic mild cognitive impairment (aMCI) as an intermediate stage. All subjects (9 AD, 16 aMCI patients, and 13 healthy controls) underwent a full neuropsychological assessment and an MRI examination at 3 Tesla, including a volumetric scan and diffusion tensor (DT)-MRI. The volumes were processed to perform a voxel-based morphometric analysis of GM and WM volume, while DT-MRI data were analyzed using tract based spatial statistics, to estimate changes in fractional anisotropy and mean diffusivity data. GM and WM volume and mean diffusivity and fractional anisotropy were compared across the three groups, and their correlation with cognitive functions was investigated. While AD presented a pattern of widespread GM atrophy, tissue loss was more subtle in patients with aMCI. WM atrophy was mainly located in the temporal lobe, but evidence of WM microscopic damage, assessed by DT-MRI, was also observable in the thalamic radiations and in the corpus callosum. Memory and executive functions correlated with either GM volume or fractional anisotropy in fronto-temporal areas. In conclusion, this study shows a comprehensive assessment of the brain tissue damage across AD evolution, providing insights on different pathophysiological mechanisms (GM atrophy, Wallerian degeneration, and brain disconnection) and their possible association with clinical aspects of cognitive decline.

Pages 161-169Mi Li, Miao Sun, Yi Liu, Jia Yu, Huan Yang, Dongsheng Fan, Dehua Chui (Communicated by Chengxin Gong)Copper Downregulates Neprilysin Activity Through Modulation of Neprilysin Degradation Abstract: Copper plays a central role in conserved processes such as respiration, and in highly specialized processes, such as protein modification. The metalloprotease neprilysin (NEP) degrades a variety of bioactive peptides and is involved in many physiological processes. However, very little is known about the regulation of NEP activity. In the current study, we focused on the effect of Cu2+ on the enzymatic activity and protein stability of NEP. Using mouse neuroblastoma N2a cells, we found that the enzymatic activity of NEP was decreased by treatment with Cu2+ in a dose- and time-dependent manner. In our investigation of the mechanism by which Cu2+ downregulates NEP enzyme activity, we found that treatment with Cu2+ caused a decrease in the level of NEP as determined by Western blotting analysis. Quantitative analysis of NEP mRNA with RT-PCR excluded the possibility that Cu2+ downregulates NEP protein at the gene transcription level. Moreover, specific proteasome inhibitors, MG132 and lactacystin, blocked the turnover of NEP, whereas inhibitors of lysosome had no significant effect, suggesting that Cu2+ induced degradation of NEP is via a proteasome pathway. Taken together, our data suggest that copper downregulates NEP activity through modulation of NEP protein degradation.

Pages 171-177Daniela Galimberti, Chiara Fenoglio, Francesca Cortini, Maria Serpente, Eliana Venturelli, Chiara Villa, Francesca Clerici, Alessandra Marcone, Luisa Benussi, Roberta Ghidoni, Salvatore Gallone, Diego Scalabrini, Ilaria Restelli, Filippo Martinelli Boneschi, Stefano Cappa, Giuliano Binetti, Claudio Mariani, Innocenzo Rainero, Maria Teresa Giordana, Nereo Bresolin, Elio ScarpiniGRN Variability Contributes to Sporadic Frontotemporal Lobar Degeneration Abstract: Mutations in progranulin gene (GRN) are responsible for familial FTLD with ubiquitin pathology (FTLD-U). However, there are controversial data regarding the contribution of GRN variability to sporadic FTLD. We carried out an association study in 265 patients who did not carry any GRN causal mutation compared with 375 age-matched controls. To this aim, four tagging Single Nucleotide Polymorphisms (SNPs) were chosen in order to get 80% power to detect an allelic association with P≤0.01. In addition, a known functional SNP (rs5848) was included. An increased frequency of the rs4792938 CC genotype in cases compared with controls was observed (17.4 versus 10.4%, P=0.01, OR: 1.81, 95%CI: 1.15-2.85). Stratifying for gender, no differences were observed for all polymorphisms. Haplotype analysis failed to detect haplotypes associated with the disease. GRN rs4792938 CC genotype represents a susceptibility factor for the development of FTLD in individuals who do not carry GRN causal mutations. This SNP is likely located in a regulatory region, thus an effect on GRN mRNA levels could be conceivable.

Pages 179-189Rui Wang, James S. Malter and Deng-Shun WangN-Acetylcysteine Prevents 4-Hydroxynonenal- and Amyloid-β-Induced Modification and Inactivation of Neprilysin in SH-SY5Y CellsAbstract: As one of the dominant amyloid-β peptide (Aβ) proteases, neprilysin (NEP) plays a crucial role in maintaining a physiologic balance between Aβ production and catabolism. We have previously shown that NEP is modified by 4-hydroxynonenal (HNE) adducts, resulting in decreased activity in the brain of AD patients and cultured cells. In order to determine whether antioxidants can rescue NEP, SH-SY5Y cells were treated with HNE or Aβ, together with N-acetylcysteine for 24 hours, prior to analysis of NEP protein levels, activity, and oxidative modifications. Intracellular NEP developed HNE adducts after 24 hours of HNE or Aβ treatment as determined by immunoprecipitation, immunoblotting, and double immunofluorescence staining. N-acetylcysteine at 10 to 100 μM alleviated HNE adduction after HNE or Aβ treatment. In keeping with previous reports, HNE-modified NEP showed decreased catalytic activity. The present study demonstrates that antioxidants can be used to spare NEP from oxidative modification, suggesting a potential mechanism underlying the neuroprotective effects of antioxidants in aging or Alzheimer’s disease.

Pages 221-228Mini-Forum Article
William R. MarkesberyNeuropathologic Alterations in Mild Cognitive Impairment: A ReviewAbstract: Mild cognitive impairment (MCI), the earliest clinically detectable phase of the trajectory toward dementia and Alzheimer’s disease (AD), catalyzed the desire for even earlier detection and prevention of AD. Although it is a clinical diagnosis, its underlying neuropathological findings are just being defined. MCI is best studied in longitudinally-followed patients in centers that are experienced in dementing disorders. In this review of the few major clinical-pathological reports of longitudinally-followed patients, it appears that most autopsied amnestic MCI (aMCI) patients are on a pathway toward AD. Neurofibrillary pathology in entorhinal cortex, hippocampus, and amygdala—not amyloid plaques—is the major substrate for aMCI and for memory decline. In addition, many MCI patients have other concomitant pathological alterations, the most common of which are strokes, but also include argyrophilic grains and Lewy bodies. These findings are not surprising because most MCI autopsied cases have been in the older (80 to 90 year) range where these findings are common. In early AD, the phase following MCI, the significant change is an increase in neurofibrillary tangles in the neocortex that correlates with an increase in Braak score and the observed clinical progression.

Pages 229-251Mini-Forum Article
Christiane Reitz and Richard MayeuxUse of Genetic Variation as Biomarkers for Mild Cognitive Impairment and Progression of Mild Cognitive Impairment to DementiaAbstract: Cognitive impairment is prevalent in the elderly. The high estimates of conversion to dementia have spurred the interest in identification of genetic risk factors associated with development of cognitive impairment and or its progression. However, despite notable achievements in human genetics over the years, in particular technological advances in gene mapping and in statistical methods that relate genetic variants to disease, to date only a small proportion of the genetic contribution to late-life cognitive impairment can be explained. A likely explanation for the difficulty in gene identification is that it is a multifactorial disorder with both genetic and environmental components, in which several genes with small effects each are likely to contribute to the quantitative traits associated with the disease. The motivation for identifying the underlying genetic risk factors elderly is clear. Not only could it shed light on disease pathogenesis, but it may also provide potential targets for effective treatment, screening, and prevention. In this article we review the current knowledge on underlying genetic variants and the usefulness of genetic variation as diagnostic tools and biomarkers. In addition, we discuss the potentials and difficulties researchers face in designing appropriate studies for gene discovery.

Pages 253-272Mini-Forum Article
Gregory A. Jicha and Sarah A. CarrConceptual Evolution in Alzheimer’s Disease: Implications for Understanding the Clinical Phenotype of Progressive Neurodegenerative DiseaseAbstract: Over the past several decades, our understanding of Alzheimer’s disease (AD) has seen an evolution from the dichotomous concept of normal versus AD in the dementia state to a more accurate and complete appreciation of AD as a progressive disorder with clinical, biological, and pathological features occurring along a continuum from normal to end-stage disease. Integrating our understanding of the relationships and interplay between the clinical, biological, and pathological features of AD may allow the identification of AD at even preclinical, completely asymptomatic stages of the disease. This review attempts to summarize the clinical stages of AD in terms of epidemiology, historical evolution of disease stage diagnoses, cognitive/neuropsychologic features, psychiatric/behavioral manifestations, and functional decline in the context of our developing understanding of the biological processes responsible for the pathogenesis of AD described in detail in the accompanying articles.

Pages 273-290Mini-Forum Article
Charles D. SmithNeuroimaging Through the Course of Alzheimer’s DiseaseAbstract: This review uses an image-indexed framework for the full course of Alzheimer’s disease (AD) to provide a perspective on recent neuroimaging findings in the literature. Modalities considered include morphometric, diffusion tensor, and functional magnetic resonance imaging, and resting/functional metabolic and amyloid-label positron emission tomography. The major focus is on the AD pre-states (normal but high AD risk and mild cognitive impairment), and transition from mild cognitive impairment to mild AD. Imaging results relevant to the conduct of future prevention and early intervention trials are emphasized.

Pages 291-300Mini-Forum Article
Juha O. Rinne and Kjell NågrenPositron Emission Tomography in At Risk Patients and in the Progression of Mild Cognitive Impairment to Alzheimer’s DiseaseAbstract: Mild cognitive impairment (MCI) is considered a transitional state between the cognitive changes of normal aging and the earliest clinical features of Alzheimer’s disease (AD). An important goal is to find features that predict which MCI patients will later convert to AD. Identification of such features will be increasingly important when treatments slowing down the progression of AD become available enabling early intervention. Brain imaging might be one possible predictor of conversion to AD. Functional imaging with positron emission tomography (PET) has shown that either normal elderly people carrying apolipoprotein E ε4 allele or people with MCI already show reduced cerebral glucose metabolism in those brain areas that are typically affected in AD. Investigations of different neurotransmitter systems might increase specificity and help in the differential diagnosis between dementing disorders. Dopamine transporter imaging to aid in the differential diagnosis between AD and dementia with Lewy bodies seems promising. Amyloid imaging is an example of “pathology specific” imaging that has great potential to enhance early detection of AD processes and to help in differential diagnosis. In the future, multi-tracer imaging or development of agents enabling imaging of other protein aggregations in neurodegenerative diseases could further help in the early and differential diagnostics and evaluation of novel treatments.

Pages 301-309Mini-Forum Article
Joshua A. Sonnen, Kathleen S. Montine, Joseph F. Quinn, John C.S. Breitner, Thomas J. MontineCerebrospinal Fluid Biomarkers in Mild Cognitive Impairment and Dementia Abstract: Given the magnitude of the public health problem of dementia in the elderly, there is a pressing need for research, development, and timely application of biomarkers that will identify latent and prodromal illness as well as dementia. Although identification of risk factors and neuroimaging measures will remain key to these efforts, this review focuses on recent progress in the discovery, validation, and standardization of cerebrospinal fluid (CSF) biomarkers, small molecules and macromolecules whose CSF concentration can aid in diagnosis at different stages of disease as well as in assessment of disease progression and response to therapeutics. A multimodal approach that brings independent information from risk factor assessment, neuroimaging, and biomarkers may soon guide physicians in the early diagnosis and management of cognitive impairment in the elderly.

Pages 311-323Mini-Forum Article
M. Paul Murphy and Harry LeVine, IIIAlzheimer’s Disease and the Amyloid-β PeptideAbstract: Alzheimer’s disease (AD) pathogenesis is widely believed to be driven by the production and deposition of the amyloid-β peptide (Aβ). For many years, investigators have been puzzled by the weak to nonexistent correlation between the amount of neuritic plaque pathology in the human brain and the degree of clinical dementia. Recent advances in our understanding of the development of amyloid pathology have helped solve this mystery. Substantial evidence now indicates that the solubility of Aβ, and the quantity of Aβ in different pools, may be more closely related to disease state. The composition of these pools of Aβ reflects different populations of amyloid deposits and has definite correlates with the clinical status of the patient. Imaging technologies, including new amyloid imaging agents based on the chemical structure of histologic dyes, are now making it possible to track amyloid pathology along with disease progression in the living patient. Interestingly, these approaches indicate that the Aβ deposited in AD is different from that found in animal models. In general, deposited Aβ is more easily cleared from the brain in animal models and does not show the same physical and biochemical characteristics as the amyloid found in AD. This raises important issues regarding the development and testing of future therapeutic agents.

Pages 325-339Mini-Forum Article
Bert C. Lynn, Jianquan Wang, William R. Markesbery, Mark A. LovellQuantitative Changes in the Mitochondrial Proteome from Subjects with Mild Cognitive Impairment, Early Stage, and Late Stage Alzheimer’s Disease Abstract: The major barrier to treating or preventing Alzheimer’s disease (AD) is its unknown etiology and pathogenesis. Although increasing evidence supports a role for mitochondrial dysfunction in the pathogenesis of AD, there have been few studies that simultaneously evaluate changes in multiple mitochondrial proteins. To evaluate changes in sites of potentially interacting mitochondrial proteins, we applied 2-dimensional liquid chromatography coupled with tandem mass spectrometry and the isotope coded affinity tag method to identify and quantify proteins in mitochondrial enriched fractions isolated from short postmortem interval temporal pole specimens from subjects with mild cognitive impairment (4 subjects pooled), early AD (4 subjects pooled), late-stage AD (8 subjects pooled) and age-matched normal control (7 subjects pooled) subjects. A total of 112 unique, non-redundant proteins were identified and quantified in common to all three stages of disease progression. Overall, patterns of protein change suggest activation of mitochondrial pathways that include proteins responsible for transport and utilization of ATP. These proteins include adenine nucleotide translocase, voltage dependent anion channels, hexokinase, and creatine kinase. Comparison of protein changes throughout the progression of AD suggests the most pronounced changes occur in early AD mitochondria.

Pages 341-353Mini-Forum Article
Rukhsana Sultana and D. Allan ButterfieldRole of Oxidative Stress in the Progression of Alzheimer’s DiseaseAbstract: Alzheimer’s disease (AD) is a progressive neurodegenerative disorder that is characterized pathologically by the presence of senile plaques, neurofibrillary tangles, and synapse loss. Increasing evidence supports a role of amyloid-β peptide (Aβ)-induced oxidative stress in the progression and pathogenesis of AD. In this review, we summarize evidence for a role of oxidative stress in the progression of AD by comparing the appearance of the same oxidized brain proteins from subjects with mild cognitive impairment (MCI), early AD (EAD), and late-stage AD, and relating these findings to the reported AD pathology. The identification of oxidized brain proteins in common in MCI, EAD, and AD brain suggest that certain key pathways are triggered and may be involved in the progression of AD. Exploring these pathways in detail may provide clues for better understanding the pathogenesis and progression of AD and also for the development of effective therapies to treat or delay this dementing disorder.

Pages 355-361Mini-Forum Article
Edith G. McGeer and Patrick L. McGeerNeuroinflammation in Alzheimer’s Disease and Mild Cognitive Impairment: A Field in Its InfancyAbstract: Neuroinflammation is a prominent feature of Alzheimer disease (AD) and other chronic neurodegenerative disorders. It exacerbates the fundamental pathology by generating a plethora of inflammatory mediators and neurotoxic compounds. Inflammatory cytokines, complement components, and toxic free radicals are among the many species that are generated. Microglia attack the pathological entities and may inadvertently injure host neurons. Recent evidence indicates that microglia can be stimulated to assume an antiinflammatory state rather than a proinflammatory state which may have therapeutic potential. Proinflammatory cytokines include IL-1, IL-6 and TNF, while antiinflammatory cytokines include IL-4 and IL-10. Complement activation is a separate process which causes extensive neuronal damage in AD through assembly of the membrane attack complex. Aggregated amyloid-β is a potent activator of human complement but not of mouse complement. This is an important difference between AD and transgenic mouse models of AD. Many so far unexplored molecules may contribute to neuroinflammation or act to inhibit it. Stable isotope labeling by amino acids in cell culture (SILAC) analysis identified 174 proteins that were upregulated by two-fold or more, and 189 that were downregulated by 2-fold or more following inflammatory stimulation of microglial-like THP-1 cells. Neurotoxicity may result from any combination of these and further exploration is clearly warranted. In addition, many small molecules may play a significant role. One example is hydrogen sulfide which appears to be an endogenous antiinflammatory agent.

Pages 363-372Mini-Forum Article
Mark A. Smith, Xiongwei Zhu, Massimo Tabaton, Gang Liu, Daniel W. McKeel, Jr., Mark L. Cohen, Xinglong Wang, Sandra L. Siedlak, Takaaki Hayashi, Masao Nakamura, Akihiko Nunomura, George Perry (Handling Editor: Jesus Avila)Increased Iron and Free Radical Generation in Preclinical Alzheimer Disease and Mild Cognitive ImpairmentAbstract: It is now established that oxidative stress is one of the earliest, if not the earliest, change that occurs in the pathogenesis of Alzheimer’s disease (AD). Consistent with this, mild cognitive impairment (MCI), the clinical precursor of AD, is also characterized by elevations in oxidative stress. Since such stress does not operate in vacuo, in this study we sought to determine whether redox-active iron, a potent source of free radicals, was elevated in MCI and preclinical AD as compared to cognitively-intact age-matched control patients. Increased iron was found at the highest levels both in the cortex and cerebellum from the pre-clinical AD/MCI cases. Interestingly, glial accumulations of redox-active iron in the cerebellum were also evident in preclinical AD patients and tend to increase as patients became progressively cognitively impaired. Our findings suggests that an imbalance in iron homeostasis is a precursor to the neurodegenerative processes leading to AD and that iron imbalance is not necessarily unique to affected regions. In fact, an understanding of iron deposition in other regions of the brain may provide insights into neuroprotective strategies. Iron deposition at the preclinical stage of AD may be useful as a diagnostic tool, using iron imaging methods, as well as a potential therapeutic target, through metal ion chelators.